Effects of an extract of khat (Catha edulis) on the toad heart

Journal of Ethno~~~olo~, Elsevier Scientific Publishers

245

18 (1986) 245-256 Ireland Ltd.

EFI;E<:‘I’S OF AN EXTRACT TOAD HEART

OF KHAT ((:AI’ZZA Z!2lliLZS)ON THE

ZOHOOR NABIL, M~STAFA SALEH”, HAMDY

~~~~WY~l

and GAMAL

ABD ALLAH”

Department of Zoology, Fwutty of Science, Suez Canal Uniuersity, ismaelia, “Department of Zoology, Faculty of Science, Al A~har University, Cairo and h3iologicaL Laboratory, National Center for ~~iologic~~ and Crim~nologiea~ Research, Cairo (eyptf (Accepted

October

14, 1986)

Summary of an extract of Catha ECG and ventricular action potential were recorded simultaneously from the surface of the heart of spinal pithed toads before and after administration of different doses of khat extract. Acute treatment resulted in a dose-dependent chronotropic effect and an increase in the amplitude of ventricular action potential. Chronic treatment on the other hand resulted in a negative chronotropic effect and a reduction in the amplitude of ventricular action potential. The observed responses are explained on the basis of the ca~cho~amine releasing effect of components of khat extract. The effects

of acute and chronic

administration

edulis on the heart of the toad were investigated.

Introduction Fresh and dried leaves and young branches of khat (Cutha edutis Forsk., Celastraceae) are widely used in parts of the Arabian Peninsula and East Africa as a stimulant. The central stimulating effects experienced by users of this plant include euphoria, enhancement and facilitation of association and increased alertness (Halbach, 1972). The pharmacological aspects of the chewing of khat have been reviewed by Kalix and Braenden (1985). Several alkaloids of the phenylalkylamine type have been isolated and identified from extracts of this plant (El Kiey et al., 1968; Karawya et al., 1968; Halbach, 1972; UN Document, 1975; Schorno and Steinegger, 1979). The stimulating activity of khat was initially attributed to the alkaloid cathine (norpseudoephedrine) (Winterfeld and Bernsman, 1960; Alles et al., 1961). More recently, however, another alkaloid cathinone has been reported to be a more potent stimulant than cathine and may possibly be 0376-8741/86/$04.55 @ 1986 Elsevier Scientific Published and Printed in Ireland

Publishers

Ireland Ltd.

246

the main active principle in C. edutis (Schorno and Steinegger. 1979; Kalix. 1980a; Zelger and Carlini, 1980). Several studies have presented evidence of chemical as well as pharmacological resemblances between cathinone and amphetamine (Halbach, 1972; UN Document, 1975; Schorno and Steinegger, 1979: Kalix. 1980a,b; 1981; WHO Advisory Group, 1980; Zelger and Carlini, 1980; 1981; Johanson and Schuster, 1981; Glennon and Liebowitz, 1982; Mereu et al.. 1983). Khat administration is known to have numerous sympathomimetic effects on both humans and animals (Le Bras and Fretillere, 1965; Raymond-Hamet. 1965; Halbach. 1972; Heacock and Forrest. 1974; WHO Advisory Group, 1980) including those affecting the ~ardi~)vas~ular system. Khat is reported to induce tachycardia and increased blood pressure in man (Halbach. 197‘2) and experimental animals (Paris and Moyse, 1957; Raymond-Hamet, 1965). Similar effects result after the injection of cathinone (WHO Advisory Group, 1964; Kohli and Goldberg. 1982). Bradycardia, palpitation and impairment of cardiovascular regulatory functions have also been reported in khat-treated animals and humans depending on the quantity of active material absorbed (Halbach, 1972). Acute and chronic administration of khat extract are reported to cause marked disturbances in the cardiac rhythm of the pithed toad (Saleh et al.. 1985). The present report provides information on the acute and chronic effects of administ,rati(~n of whole khat extract on the heart of the toad. Experiment.al

EIxiractior~ arid do.ses Fresh leaves of C’. edulis obtained from plants grown in the Botanical Gardens of the Ministry of Agriculture, Qanater, Egypt were used. The* crushed leaves were extracted according to the method of Alles et al f 1961) in 1: 3 (v/v) chloroform/diethylether mixture and the solvents evaporated under reduced pressure. The dried extract represented 0. f 1% of the dry weight of khat leaves. For injection, the extract thus obtained was suspended in a 1: 1 mixture of Tween-80 (polysorbate-80) and amphibian Ringer’s solution immediately prior to injection. The suspension was injected in the lymph sac of the toad in a volume of 0.01 ml/g body wt containing the calculated dose of the extract. Three doses of khat extract, 2576, 50%. and 100% of the median lethal dose of 0.81 mglg body wt (Saleh et al., 1985) were used in the acute experiments. In chronic experiments, a dose of 10% of the median lethal dose was injected daily for a total period of 15 days. Control animals underwent exactly the same treatment as experimental animals but were injected with Tween-80famphibian Ringer’s solution mixture at the same volume and rate as the experimental animals.

247

An irnals and surgical

preparations

Adult male and female toads (Bufo regularis Reuss) 35-40 g body wt were used. Animals subjected to the acute experiments were collected from the field on the morning of the experiment. Those used for the chronic experiments were kept in a glass aquarium containing a thin layer of moist soil and fed on insect larvae throughout the experimental period. All experimental work was carried out during the spring and summer, the normal activity season for this toad in Egypt, and at an ambient temperature of 24 f 4°C. Forty animals, divided into four equal groups were used for the acute experiments. One group served as a control and received TweenHO/amphibian Ringer’s solution in a volume of 0.01 ml/g body wt, and each of the three treated groups received one of each of the three doses. Forty five animals were used in the chronic experiments, 15 control and 30 treated. A group of 5 control and 10 treated animals were tested 5, 10 and 15 days after the onset of the treatment. For exposing the heart. the toad was spinal pithed, a mid-ventral incision was made in the skin, and the underlying muscles were carefully separated by blunt dissection. The pericardium was then carefully removed exposing the myocardium. A few drops of amphibian Ringer’s solution were applied to the heart to keep it from drying.
248

heart using two pieces of cotton thread, about 5 cm in length each, saturated with Ringer’s solution. The end of one thread was placed on the right atrium and the end of the other on the apex of the ventricle. The cotton threads, which were always kept saturated with Ringer’s solution, allowed free movement of the heart while maintaining good electrical contact. ECG signals were picked up through two silver wires in c~ontnct with the two cotton threads. amplified and recorded on the chart recorder and monitored on an oscilloscope. In the acute experiments, a recording was carried out. before the injection. Then the animal was injected and recording wits carried out 5. 15, 30, 45. 60, 90 and 120 min following the injection. In chronic experiments, the animal was injected wit.h its daily dose of khat extrar-t and action potential and ECG were recorded 30 min after the injection. Differences among groups of treated animals and bet.ween each group and the control group were assessed by analysis of variance. St~~tisti~~~l significance was reached when P ‘ 0.05. Results Typical monophasic ventricular action potentiais and simultaneously recorded electrocardiograms of the normal toad are shown in Fig. 1. Thestk records are comparable to those recorded from related arnphibi~~r~specicbs using similar techniques (Hoffman, 1959; Kanno. 1962) Ac:ute eflPcts art heart rate Acute administration of khat extract resulted in an increase in heart. rate which appeared immediately after injection (Fig. 2a). The smallest, dose produced the largest increase in heart rate and this increase was maintained throughout the entire recording period of 2 h. With the largest

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Fig. 2. meets of three doses of 202, 405 and 810 cLg/g body wt of khat extract on heart rate (a) and T-P (b) and P-R intervals (c) of ECG of spinal pithed toads. Control values (mean + S.E.M.) were: heart rate, 60.0 + 1.0 beatslmin; T-P interval, 341.8 16.2 ms; and P-R interval, 245.1*4.8ms; N=80.

250

dose, the increase in heart rate was less profound and more gradual than the two smaller doses but remained significantly higher than the normal rate. Analysis of variance showed that the heart rate after extract 0.01). It also injection was significantly higher than the-control value (P showed that the responses to different doses were significantly different (P 0.01). Analysis of the ECG components showed that the observed tachyc.ardia was associated with a reduction in the T-P interval. whic.11 represents the

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Fig. 3. Effects of three doses of 202, 405 and 810 pg/g body wt of khat extract on amplitude (a) and duration (b) of ventricular action potential of spinal pithed toads. Control values (mean i S.E.M .) for amplitude and duration of ventricular action potential were 3 1 56 I 1.40 mV and 45.38 i 0.71 ms, respectively. N z 80.

251

pause between the successive beats (Fig. 2b). It also showed a reduction in the P-R interval, which represents the time required for atrioventricular conduction (Fig. 2~). Analysis of variance showed that the changes in both parameters were statistically significant (P .’ 0.01) following each of the three doses of the extract. This test also showed that the differences between the responses to different doses were statistically significant in case of the T-P interval (P. 0.01) but not in the case of the P-R interval. Acute

effects on ventricular

action

potential

Figure 3a shows the effects of acute doses of khat extract on the amplitude of the ventricular action potential. Analysis of variance showed that this increase was statistically significant (P -- 0.01) for each of the three doses of khat extract. This analysis also showed that the responses to the three doses were significantly different (P .’ 0.05). Figure 3b shows the changes in the duration of the ventricular action potential following the injection of three doses of khat extract. According to the analysis of variance, the duration of ventricular action potential

0

25

50

100

DOSEi% OF tDSO’ Fig. 4. A comparison between the mean effects of doses of khat extract on heart rate (0). T-P (Cl) and P-R (0) intervals of ECG and amplitude (e) and duration (v) of ventricular action potential of spinal pithed toads. Control values (mean f S.E.M.) were: HR, 60.0 f l.Obeats/min; T-P, 341.8*S.E.M. 6msec; P-R, 245.1k4.8 ms; amplitude, 31.56* 1.40mV; duration, 45.38 f 0.71 ms; N = 80.

252

was not significantly affected by the two larger doses of khat extract. With the smaller dose, the duration was initially not significantly different from that of the control; however, 90 and 120 min after the injection it dropped to a value significantly (P ‘_ 0.05) lower than the control value. The responses to the three doses were not significantly different from each other according to analysis of variance. In general, the acute effects of administration of khat extract appeared to be dose-dependent. Figure 4 shows dose-response curves of the effects of

DURATION

OF TREATMENT

‘DAYS]

Fig. 5. EXects of daily administration of a dose of 81 kg/g body wt of khat extract on heart rate (0) and T-P (0) and P-R (V) intervals of the ECG and amplitude (0) and duration (W of ventricular action potential. Control values (mean * S.E.M.) were: HR, 64.2 i 2.3 beatslmin: T-P, 166.0 f 13.1 ms; P-R, 228.8 + 3 9 ms; amplitude, 54.15 * 2 22 mV; duration, 32.45 i 1.61 ms; N = 40.

253

three different doses of khat extract on heart rate, T-P and P-R intervals of the ECG and the amplitude and duration of ventricular action potential. In cases of the heart rate, P-T interval and amplitude of ventricular action potential, the smallest dose produced the largest response (P ~‘0.01). In the case of the P-R interval and the duration of the ventricular action potential, the responses to the three doses were not significantly different. C’hron ic experiments

Chronic treatment with daily doses of 81 Fglg of khat extract resulted in a significant (P J 0.01) drop in the heart rate that progressed throughout the 15 days of treatment (Fig. 5). While this treatment resulted in an initial drop in the P-R interval (P c 0.01) after 5 days of the treatment, the P-R interval lo-15 days after the treatment was not significantly different from that of the control animals (Fig. 5). The T-P interval, however, showed a significant (P .: 0.01) increase as the treatment duration increased (Fig. 5). Chronic treatment with khat extract resulted in a significant (P < 0.05) decrease in the amplitude of action potential following 5-10 days of treatment. This increase became insignificant however, after 15 days of treatment (Fig. 5). The duration of action potential, on the other hand, showed a significant (P c 0.01) progressive increase reaching a maximum after 15 days of the treatment (Fig. 5). Discussion The results presented above agree well with the known sympathomimetic effects of khat extracts (Le Bras and Fretillere, 1965; WHO Advisory Group, 1980; Zelger and Carlini, 1980; Valterio and Kalix, 1982). These effects have been attributed mostly to the alkaloid (-)cathinone which closely resembles amphetamine in both structure and pharmacological effects (Winterfeld and Bernsman, 1960; Halbach, 1972; Kalix, 1980a, 1980b, 1981; Kohli and Goldberg, 1982; Glennon and Liebowitz, 1982; Valterio and Kalix, 1982). As in the case of amphetamine (Axelord et al., 1962; Schumann and Phillippu, 1962), most of the pharmacological effects of cathinone can be explained on the basis of its facilitation of the release of catecholamines from intracellular storage sites (WHO Advisory Group, 1980; Kalix, 1980a,b, 1981). The observed positive chronotropic effect of khat extract supports previous reports of similar effects after administration of khat (Le Bras and Fretillere, 1965; Halbach, 1972), a number of its components (Brucke, 1941; WHO Advisory Group, 1980; Kohl and Goldberg, 1982) and amphetamine (Simpson, 1974; WHO Advisory Group, 1964; Kohl and Goldberg, 1982). Furthermore, our data show that the increase in heart rate is the result of an increased firing rate of the sinus node cells as

254

reflected in the shortened T-P interval of the otherwise normal EC<;. The reduction of the P-R interval, which signifies a shorter atrioventricular conduction delay, is another manifestation of the overall excitation of the heart. Catecholamines are known to induce similar effects on the myogenic. hearts of the vertebrates. These transmitters, by increasing the rate of depolarization of the cardiac pacemaker and conduction speed through the atrioventricular junction, produce their well documented chronotropic effect (Ueda et al., 1964; Urthaler and James. 1973, 1976: IJrthaler et al., 1973). Similar effects of amphetamine and its derivatives are mediated through the release of catecholamines from intracellular stores (Simpson, 1974). The increased amplitude of the ventricular action potential observed following administration of khat extract is also consistent with the sympathomimetic nature of its active components. The amplitude of ventricular action potential is reported to increase in response to adrenergic neurotransmitters and amphetamine treatment (Hoffman et a1 1959; Levy and Zieske. 1969; Urabe, 1982). Amphetamine, as well as a numher of khat components are reported to have positive inotropic effects on isolated atria of guinea pigs (WHO Advisory Group, 1980). The time required for ventricular depolarization and repolarization is represented by the duration of the ventricular action potential. This time interval was not significantly affected by the administration of the two larger doses of the extract. The small but significant decrease in this duration, which occurred at 90 and 120 min after administration of the 202 kg/g dose may indicate a reduced ventricular refractory period and higher velocity of ventricular contraction. Randall (1976) similarly found that ventricular refractoriness, which is directly related to the duration of depolarization plateau, clearly shortened by sympathetic stimulation. The results also show that the smallest dose of the khat extract had the greatest effect on heart rate, T-P interval and amplitude of ventricular action potential, while the effects become smaller as the dose has increased. This somewhat resembles the dose-response relationship of cathinone hypermotility which is reported to be similar to that of amphetamine-induced hypermotility (Valterio and Kalix, 1982). This further supports an amphetamine-like mechanism of action for khat which involves the release of catecholamines from intracellular storage sites (Axelord et al., 1962; Schumann and Phillippu, 1962; Kalix, 1980a,b, 1981; WHO Advisory Group, 1980). Contrary to the situation with the acute treatment, chronic treatment with khat extract produced a general depression in the heart functions. Heart rate gradually declined throughout the study period as a result of the slower firing rate of the cardiac pacemaker leading to the observed increase in the T-P interval. This depression of cardiac function was also evident in the increased duration and the decreased amplitude of the ventricular action potential. Atrio-ventricular conduction speed did not

appear to change except during the earlier period of the treatment as the P-R interval showed a significant reduction only after 5 days of the treatment. The data presented here, therefore, support reports that khat administration has pharmacological effects similar to those obtained by administration of amphetamines and its derivatives. The data also show that the administration of this widely consumed stimulant has drastic effects on the heart, particularly when larger doses are taken on a daily basis. i\cknowledgments

We are grateful construction

and

to Dr. Wahib A. Mohamed for his help in the of the flexible suction electrode system.

operation

References Alles, G.A.,

Fairchild,

M.D. and Jensen,

M. (1961) Chemical

pharmacology

of Catha edulis.

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